Microbiological Analysis of Drinking Water

Test for Coliforms and E.coli

Group 3 3-A Biochemistry
Cua, G.A; Diego, J.L; Divina, T.R; Dulatre, E.A

Abstract
Water is one of the most important substances on earth. Drinking water must be free of disease-causing organisms called pathogens.
Pathogens can be viruses, protozoa or bacteria. For this experiment three samples of drinking water from UST were taken from the
following locations: the drinking fountain beside the female washroom on the first floor of the Quadri Pavilion , the faculty room, and the
fountain in front of lab 19 located at the Main Building of the Faculty of Pharmacy. Moreover, the sources were then put through the
process of serial dilution , yielding the dilution factors of 10-1,10-2 and 10-3. Lastly, each dilution had three test tubes . Therefore, there
were three samples, each having three different dilutions , and three trials per dilution. In conclusion, each of the water samples
gathered were proven to be unsafe for consumption due to it having an MPN number greater than 1 in the test for E. coli, water should
be free of Total Coliforms to be considered totally safe to drink. Lastly, the main objectives of this experiment were the following: to
determine the potential presence of coliforms and E. coli in water , to skillfully perform multitube technique in technique in testing total
coliforms.

Introduction
Water is one of the most important substances on earth. All plants and
animals must have water to survive. If there was no water there would be no
life on earth.[1] Moreover , an average adult body contains 42 liters of water
and with just a small loss of 2.7 liters he or she can suffer from dehydration,
displaying symptoms of irritability, fatigue, nervousness, dizziness, weakness,
headaches and consequently reach a state of pathology. [2] Also, it serves as a
lubricant for digestion, regulation of body temperature, removes harmful
toxins from the body, and transports nutrients to the body.[3]
Drinking water must be free of disease-causing organisms called pathogens.
Pathogens can be viruses, protozoa or bacteria. Waterborne pathogens cause
diseases such as hepatitis, giardiasis, and dysentery. To actually test water for
specific harmful viruses, protozoa and bacteria is very time consuming and
expensive. In addition, not all water laboratories are equipped and approved
to do the testing required. Therefore, testing water for specific organisms is
limited to investigating specific waterborne disease outbreaks. Coliform
bacteria are used as water quality indicators for two main reasons: Coliforms
may be associated with the sources of pathogens contaminating water,
analysis of drinking water for coliforms is relatively simple, economical and
efficient.[5]
Escherichia coli which is in the large intestine of virtually all people has been
used as the indicator of human fecal contamination of water and food. Tests
for the presence of this organism (and
closely related types generally known as coliforms .[7] Coliforms are a broad
class of bacteria found in our environment, including the feces of man and
other warm-blooded animals. The presence of coliform bacteria in drinking
water may indicate a possible presence of harmful, disease-causing
organisms.[5]
The MPN method is a well-established and fully documented method that
utilizes the multitube technique in order to estimate the number of
viable microorganisms in product in which the microorganisms are randomly
distributed. Its principal uses are in the food and water industries where it is
used with liquid, powdered and semi-solid products or raw materials. The
method is particularly appropriate for product having bio burden of low mean
number. [7]
Lastly, the main objectives of this experiment were the
following: to determine the potential presence of coliforms and E. coli in
water , to skillfully perform multitube technique in technique in testing total
coliforms.

Methodology

prepare necessary
media for Presumptive
Test , Test for E. coli,
Test for Coliforms and
store

Gather water samples using

a sterile container and
inoculate serially diluted
samples to pre-prepared
media and wait for 24-48
hours

Determine
results or
growth
and record
results

Results and Discussion

For this experiment three samples of drinking water from UST were taken
from the following locations: the drinking fountain beside the female
washroom on the first floor of the Quadri Pavilion , the faculty room, and
the fountain in front of lab 19 located at the Main Building of the Faculty
of Pharmacy. Moreover, the sources were then put through the process of
serial dilution , yielding the dilution factors of 10-1,10-2 and 10-3. Lastly,
each dilution had three test tubes . Therefore, there were three samples,
each having three different dilutions , and three trials per dilution.
Table 1.1 Results obtained from the Presumption Test with MPN

Accordingly, a 3-tube MPN table was used in order to determine the MPN
value of each sample for the tests. For the test for E. coli only the growth has
to be positive to be counted as a part of the MPN. Therefore , this yielded a
result of >24 for the Quadri Pavilion drinking fountain ,11 for the faculty
room and 2.4 for the lab 19 drinking fountain . (See source [4] for the 3tube MPN table)
The results showed that P. aeruginosa, P. mirabilis may exist in the Quadi
Pavilion drinking fountain, P. aeruginosa may exist in the faculty room
water , and P. aeruginosa , E. faecalis, E. aerogenes , B. subtilis may exist in
the lab 19 drinking fountain water.
Table 1.2 Results obtained from the Confirmation Test with MPN

For the Test for Coliforms, the Brilliant Green Bile broth was used in order
detect or confirm the presence of members of the coli-aerogenes group; the
brilliant green content suppresses anaerobic lactose fermenters. Also, a
Durham tube is provided to detect the production of gas.
Accordingly, a 3-tube MPN table was used in order to determine the MPN
value of each sample for the tests. For the test for coliforms both growth
and gas production have to be positive to be counted as a part of the MPN.
Therefore, this yielded a result of 0.094 for the Quadri Pavilion drinking
fountain, less than 0.03 for both the faculty room and the lab 19 drinking
fountain.
The results showed that E. faecalis possibly thrives in all the sample
sources.
Table 1.5 Results obtained from the Completed Test for E. coli

For the Completed Test for E. coli , Eosin methylene blue agar was used in
order to confirm the presence of E. coli in the water. It is used in water
quality tests to distinguish coliforms and fecal coliforms that signal possible
pathogenic microorganism contamination in water samples, contains
sucrose and lactose which serves as fermentable carbohydrate source to
encourage the growth of fecal coliforms and provide a means of
differentiating them.
There are several different types of growths that may be observed in the
agar. Bacteria that ferments lactose show up as colored colonies, those
which do not ferment lactose show up colorless colonies, Escherichia
coli shows up as a green metallic sheen with a dark center, Aerobacter
aerogenescolonies show up as brown centered colonies , and nonlactosefermenting gram-negative bacteria show up as pink colorations.

Conclusion
In conclusion, each of the water samples gathered were proven to be unsafe for
consumption due to it having an MPN number greater than 1 in the test for E.
coli, water should be free of Total Coliforms to be considered totally safe to
drink

References
1.

2.
3.
4.
5.
6.

For the Presumption Test, due to the unavailability of the Mac Conkey
broth the EC broth was utilized for both this test and the Test for E. coli.
The EC broth is is used in the detection of fecal coliform bacteria as an
indication of fecal pollution of food and water from sources other than
potable water. A Durham tube is provided to detect the production of gas.

7.

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November
)
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